JP2007023051A - Particle of bioactive agent and method for manufacture thereof - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a delivery system for oral administration or the like of a poorly water-soluble biologically active agent. <P>SOLUTION: The poorly water-soluble biologically active agent is melted; a dispersing agent is heated; the biologically active agent is mixed with the dispersing agent in the presence of a dispersion stabilizer such as a phospholipid, a sphingolipid, a glycosphingolipid, a salt of bile acid, a fatty acid, an aliphatic alcohol, and an ester of a fatty acid and an aliphatic alcohol; and the dispersion is emulsified by high-pressure homogenization, and allowed to be stood to be cooled until solid particles are formed by recrystallization. As a result, the suspension of the particles (PBA) of the biologically active agent is prepared. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

  The present invention relates to a suspension of particles of a biodegradable lipid, preferably triglyceride, which is solid at room temperature, which can be used as a carrier for poorly water-soluble drugs or other bioactive agents, and bioactive agents such as pharmaceuticals, insecticides, In addition to suspensions of particles composed of fungicides, pesticides, herbicides and fertilizers, etc., they also relate to lyophilized products thereof. Either system can be prepared by melt emulsification.

The properties of solid lipid particles (SLP) include biodegradability, avoidance of toxicologically active residues from the production process, enhanced physicochemical stability with respect to coalescence and chemical leakage, modified surface properties This includes controlled release of incorporated substances and modified biodistribution. The particles can be prepared by a melt mass emulsification method that produces small droplets that form crystalline anisometrical particles upon cooling. The non-equal axis particles are micron and submicron in size, mainly in the range of 50-500 nm. The aforementioned suspensions are present because the solid biodegradable matrix exists primarily as β-polymorphic modifications (eg, β ′, β ₁ , β ₂ ) rather than in an amorphous or α-crystalline state, There are several advantages over other drug carrier systems.

  Preparation of micron and submicron particles (PBA) consisting of poorly water-soluble bioactive agents by melt material emulsification provides a new method for reducing particle size and / or modifying the surface properties of powdered materials. It can also be done by cheap methods and using only physiologically acceptable additives. This particle suspension is a product that is easy to handle from the viewpoint of security. The particles of bioactive agent define the modified biodistribution and bioavailability of the formulated drug or other bioactive substance, which is the drug or other bioactive It suggests that the degree and rate of dissolution and absorption of substances, circulation time, site of action and excretion mode are modified. Due to particle size reduction below the micrometer range, particles made from poorly water-soluble drugs can be directly administered intravenously without the need for a carrier vehicle.

  The present invention falls within the field of dosage forms and delivery systems for pharmaceuticals, vaccines and other bioactive agents such as insecticides, fungicides, pesticides, herbicides and fertilizers. More specifically, the present invention provides for the preparation of suspensions of colloidal solid lipid particles (SLP) and bioactive agent micron and submicron particles (PBA) whose lipid matrix is in a stable polymorph. In addition to such parenteral administration, preferably mainly poorly water-soluble biologically active substances, especially pharmaceuticals, are also used for oral, nasal, pulmonary, rectal, transdermal and buccal administration. It relates to the use of suspensions or their lyophilizates as delivery systems and their use in cosmetics, food and agricultural products. These suspension systems provide controlled release of the incorporated or constituent material, as well as modified biodistribution and bioavailability of the incorporated or constituent drug. However, this suggests that the degree and rate of dissolution and absorption of the drug, circulation time, site of action and excretion mode are modified.

  Parenteral, especially intravenous, administration of water insoluble or poorly water soluble materials such as pharmaceuticals or other biological materials is often a problem for formulators. The smallest capillary diameter is only a few microns, so intravenous administration of larger particles will occlude the capillary. However, solid drug substances are usually disrupted by milling and grinding, thereby producing particles in the millimeter to micrometer size range, which are too large to be directly injected as an aqueous suspension. As a result, intravenous systems containing suspended particles of water-insoluble drugs are not commercially available due to the risk of embolism. Further particle size reduction is expensive by conventional methods and is ineffective or even impossible. In addition, reducing solids to submicron sized powders creates significant problems in handling these dried products, such as increased risk of dust explosions and cross-contamination problems in the factory environment. Furthermore, such systems pose a health hazard to those exposed to inhaled and absorbed potentially bioactive substances. To date, the only possibility to administer poorly water-soluble substances by the intravenous route is the use of cosolvents or the development of carrier systems that incorporate such substances into vehicles with hydrophilic surfaces.

  The basic requirements of an ideal pharmaceutical carrier system include biodegradability, non-toxicity, and non-immunogenicity. In addition, the carrier should be suitable for the intended route of administration, for example with respect to particle size. For example, controlled release of incorporated bioactive substances is often desirable when constant serum levels are to be maintained over a long period of time, or when the medication exhibits a low therapeutic index.

  Furthermore, carrier systems can be used to increase the half-life of certain substances that are unstable because they rapidly degrade enzymatically or hydrolytically in a biological environment. On the other hand, incorporation of a medicament into a carrier material also provides an opportunity to protect the host from the medicament in the case of non-selective toxic substances such as anti-tumor agents.

  In many cases, pharmaceutical carrier systems are developed with the purpose of delivering drugs to site-specific targets while bypassing uptake by the reticuloendothelial system (RES). The rationality of such drug targeting is to increase the therapeutic efficacy of the drug because the dose can be reduced because the drug concentration increases at the target site and simultaneously decreases at the non-target site. That is, for example, the toxicity of a medicine such as an anticancer agent can be reduced, and the side effects are reduced.

  Prerequisites for successful site-specific delivery include a certain selectivity of the carrier system for the target tissue and accessibility of the desired target site. Targeting by the intravenous route generally leads to avoidance or at least reduction of carrier uptake by RES, unless direct targeting to RES cells is desired. The clearance of colloidal particles by RES has been described as dependent on particle size and particle surface properties such as surface charge and surface hydrophobicity. In general, small particles have a slower clearance from the bloodstream than large particles, whereas charged particles are taken up more rapidly than hydrophilic uncharged particles. Because of these facts, surface property modification and particle size reduction are taken as approaches to drug targeting.

  In addition, small particle sizes are required to target drugs to the extravascular location. This is because extravasation is possible only through receptor-mediated uptake by phagocytosis / drinking, or when there is a window in the endothelial wall. Such windows can be seen, for example, in the sinusoids of the liver, spleen and bone marrow, and show diameters up to about 150 nm.

  From a manufacturing point of view, an ideal pharmaceutical carrier system should be reproducible and easy to prepare, preferably at a low production cost, in a manner that is easy to handle. The formulation should exhibit sufficient stability during preparation and storage.

  In recent years, some colloidal systems have been of particular interest due to the possibility of being accepted as pharmaceutical carriers, among which are liposomes, lipid emulsions, microspheres and nanoparticles. However, since all of these systems have a certain number of drawbacks, so far no breakthrough has been opened for any such system to be used as a widespread commercially available pharmaceutical carrier.

  Pharmaceutical carrier systems in the micrometer size range are represented by microspheres composed of a solid polymer matrix and microcapsules in which the liquid or solid phase is surrounded and encapsulated by a polymer film. Nanoparticles, like microspheres, consist of a solid polymer matrix. However, their average particle size is in the nanometer range. Both micro- and nanoparticles are generally prepared by emulsion polymerization or solvent evaporation methods. Because of these production methods, micro- and nanoparticles are at risk of contamination from residues from the production process, for example organic solvents such as chlorinated hydrocarbons as well as toxic monomers, surfactants and crosslinkers. The result can cause toxicological problems. In addition, some polymeric materials, such as polylactic acid and polylactic acid-glycolic acid, degrade very slowly in vivo, resulting in polymer accumulation with deleterious side effects when administered in multiple doses. Other polymers, such as polyalkylcyanoacrylates, release toxic formaldehyde when decomposed in the body.

  Lipid based pharmaceutical carrier systems for parenteral administration are liposomes and submicron lipid emulsions. While such systems are composed solely of physiological components and thus alleviate toxicological problems, there are many disadvantages associated with these lipid carriers.

  Liposomes are spherical colloidal structures in which the internal aqueous phase is surrounded by one or more phospholipid bilayers. The possibility of using liposomes as a drug delivery system is, among others, US Patent No. 3,993,754 (issued November 12, 1976 to Rahmann and Cerny) and No. 4,235,871 (November 25, 1980 to Papahadjopoulos and Szoka). No. 4,356,167 (issued on October 26, 1982 to L. Kelly). The main drawbacks of conventional liposomes are instability during storage, low production reproducibility, low capture efficiency and leakage of medication.

  According to the definition of IUPAC, a liquid or liquid crystal is dispersed in a liquid in an emulsion. Parenteral lipid emulsions are composed of liquid oil droplets, primarily in the submicron size range, which are dispersed in the aqueous phase and stabilized by an interfacial film of emulsifier. A typical formulation is disclosed in Japanese Patent No. 55,476 / 79 issued on May 7, 1979 to Okamoto, Tsuda and Yokoyama. The preparation of drug-containing lipid emulsions is described in WO 91/02517 issued March 7, 1991 to Davis and Washington. The ease of drug uptake of these lipid emulsions is relatively high due to the mobility of the drug molecules within the internal oil phase. This is because the diffusing molecule can easily protrude into the emulsifier film, causing instability leading to coalescence. Furthermore, the potential for sustained drug release is limited due to the relatively rapid release of the incorporated drug from the lipid emulsion.

  Fountain et al (US Patent No. 4,610,868 issued September 9, 1986) is described as a spherical structure of a hydrophobic compound having a diameter of about 500 nm to about 100,000 nm and an amphiphatic compound. A lipid matrix carrier was developed. The hydrophobic compound may be liquid or solid. However, the preparation method uses an organic solvent, and therefore involves the problem of complete solvent removal.

  The so-called lipospheres disclosed by Domb et al (US Patent Application No. 435,546 filed November 13, 1989; International Application No. PCT / US90 / 06519 filed November 8, 1990) are surrounded by a phospholipid layer. It is described as a solid, water-insoluble microsphere suspension consisting of a solid hydrophobic core. Lipospheres are claimed to provide sustained release of capture substances controlled by the phospholipid layer. They can be prepared by melt or solvent methods, but the latter creates toxicological problems if the solvent is not completely removed.

  A sustained release composition suitable for parenteral administration of fat or wax and biologically active protein, peptide or polypeptide to animals is US Patent Application No. 895,608 filed on August 11, 1986 by Staber, Fishbein and Cady. (EP-A-0 257 368). Since these systems are prepared by spray drying and are composed of spherical particles in the micrometer size range up to 1,000 microns, intravenous administration is not possible.

  Problems associated with formulation of water-insoluble or poorly water-soluble substances are not limited to parenteral routes of administration. That is, oral bioavailability of drugs is related to their solubility in the gastrointestinal tract (GIT), and generally poorly water-soluble drugs are found to have low bioavailability. Furthermore, drug dissolution in GIT is affected by their wettability. Since materials with non-polar surfaces do not wet much in the medium, their dissolution rate is very slow.

  Eldem et al (Pharm. Res. 8, 1991, 47-54) prepared lipid micropellets by spray drying and spray congealing in an attempt to improve the superabsorption of lipophilic drugs. These micropellets are described as spherical particles having a smooth surface. However, lipids exist in unstable polymorphs and product properties always change because polymorphic phase transitions occur during storage (T. Eldem et al, Pharm. Res. 8, 1991, 178-184). Therefore, a constant quality cannot be guaranteed.

  Lipid nanopellets for oral administration of drugs with poor bioavailability are disclosed in EP 0 167 825 (August 8, 1990, P. Speiser). These nanopellets are drug-loaded fat particles that are solid at room temperature and small enough to undergo persorption. Persorption is the transport of whole particles through the intestinal mucosa to the lymph and blood compartments. Lipid nanopellets are prepared by emulsifying molten lipids in an aqueous phase with high speed stirring. After cooling to room temperature, the pellets are dispersed by sonication.

  Considering the limitations of traditional pharmaceutical carriers such as liposomes, lipid emulsions, nanoparticles and microspheres outlined above, avoid the disadvantages of traditional systems, especially with regard to modification of preparation, stability, toxicity and biodistribution Thus, there is a clear need for carrier systems for the controlled delivery of poorly water-soluble bioactive substances.

  The present invention provides a new type of carrier system characterized as non-spherical particles composed of crystalline lipids, preferably triglycerides, and physiologically acceptable additives, and methods for their production. These carriers are primarily parenteral, but also provide controlled delivery of poorly water-soluble substances, such as drugs or other biological substances, by nasal, pulmonary, rectal, dermal and buccal routes of administration. Hereinafter, it is referred to as solid lipid particles (SLP).

SLPs are characterized as lipid particles in the solid physical state that are micro- and predominantly in the nanometer size range. The particle shape is primarily non-axial, but this means that the lipids that form the matrix exist in β-polymorphic form (eg β ′, β ₁ , β ₂ ) and do not exist in an amorphous or α-crystalline state Is the result of Properties of SLP include: (1) biodegradable and non-toxic; (2) poorly water-soluble substance uptake capacity; (3) improved chemical and physical stability; (4) drying Possibility of preparing storage formulations; (5) Control of release characteristics of incorporated substances; and (6) Modified surface characteristics. As a result of these properties, SLP overcomes many of the problems encountered with conventional pharmaceutical carrier systems.

The present invention is believed to provide the following advantages derived from the aforementioned SLP features:
(1) SLP can be prepared with only biodegradable and pharmacologically acceptable compounds and is therefore non-toxic. Furthermore, the preparation of SLP avoids the use of organic solvents or any other potentially toxic additive, thus avoiding residual impurities in the product.
(2) SLP has a higher chemical stability than conventional lipid emulsions based on liquid triglyceride oil due to the low degree of unsaturated fatty acids in solid triglycerides. In addition, SLP exhibits better physical stability because of the solid nature of the lipid matrix, which is more resistant to coalescence than flowable emulsion droplets as expected. Furthermore, the lipid matrix exists in stable β-polymorphs (eg β ′, β ₁ , β ₂ ). That is, product properties do not change significantly during long-term storage due to polymorphic deformation.
(3) The suspension of SLP can be lyophilized, resulting in a moisture-free storage system with good long-term stability. The lyophilized powder can be redispersed in water, buffer or amino acid or carbohydrate solutions and other infusion solutions, or processed into other pharmaceutical formulations, just prior to use.

(4) Because of their lipophilicity, SLP is suitable for solubilization of lipophilic and poorly water-soluble substances by entrapment in a lipid matrix. SLP is considered less sensitive to the uptake of pharmaceuticals or other bioactive substances because of its solid nature compared to lipid emulsions. Drugs or other bioactive substances that diffuse into the emulsifier film or recrystallize near the surface disrupt the emulsion droplet stabilization film and increase the risk of film breakage with coalescence. In contrast, film elasticity and film viscosity are less important in the case of solid suspension particles. Because they cannot coalesce because of the rigidity of the lipid.
(5) The drug release from the lipid carrier can be controlled by, for example, the composition of the lipid matrix, the choice of stabilizer and the particle size of the SLP. Drug leakage is hindered by limited drug diffusion due to the solid state of the carrier.
(6) Drugs or bioactive substances that exhibit a short half-life due to enzyme or hydrolytic degradation can be protected from rapid degradation by incorporation into a lipid carrier. This is because the hydrophobic matrix hinders access to water for the pharmaceuticals incorporated not only during storage but also in body fluids.

(7) The bioavailability of such substances can be increased by incorporating drugs or other bioactive substances with low bioavailability due to poor solubility in the gastrointestinal tract (GIT) into the SLP. This is because they are solubilized in the biodegradable lipid matrix and therefore exist in a dissolved state.
(8) Since the shape of SLP is non-equal axis, its specific surface area is larger than that of isotropic spherical particles. Substances with low oral bioavailability are absorbed rapidly and highly in GIT when incorporated into non-equal axis SLPs due to the larger surface area of SLPs than in equal volume spherical lipid particles. obtain. This is because the potential site of action for lipolytic enzymes is large.
(9) The surface properties of SLP can be modified by changing lipid composition, using different stabilizers, exchanging surfactants and / or adsorbing polymeric compounds. This modification of surface properties creates the possibility of modifying the biodistribution of the carrier and the incorporated material. In the case of intravenous administration, this means that uptake is modified by RES, resulting in the possibility of drug targeting.
(10) Since the SLP particle size is submicron, there is no risk of embolism due to parenteral administration. Since SLPs can be prepared to a particle size of about 50 nm, they have the opportunity to overflow through the window of the endothelial wall. Thereby, the medicament can be targeted to an extravascular location, such as the bone marrow.

  In addition, the present invention provides a new type of delivery system for parenteral, oral, nasal, pulmonary, rectal, transdermal and buccal administration of pharmaceuticals or other bioactive substances, and methods for their production. . These formulations are suspensions of particles formed by bioactive materials having modified surface properties and / or reduced particle size compared to powdered materials, hereinafter referred to as bioactive agent particles (PBA). . The preparation of PBA can avoid the use of any toxicologically active additives such as organic solvents or toxic monomers and can be carried out in an easy to handle manner.

PBA can be used in the following application areas:
a) as a parenteral delivery system that does not require a carrier vehicle, wherein the biodistribution for poorly water-soluble bioactive substances is modified;
b) as a delivery system according to a) for oral, nasal, pulmonary, rectal, transdermal and buccal administration;
c) as a formulation for oral administration of drugs with poor bioavailability due to low dissolution rate in the gastrointestinal tract;
d) as a delivery system for use in agricultural applications;
e) Lyophilized products of formulations a) to d) as reconstitutable powders with enhanced storage stability.

Due to the special features of the present invention, PBA is believed to provide the following advantages over conventional pharmaceutical delivery systems:
1) Compositions of micro- and sub-micron particles of poorly water-soluble drugs or other bioactive substances do not require a carrier system for their parenteral administration, so conventional pharmaceutical carriers such as liposomes, lipid milk The disadvantages of suspending agents, nanoparticles and microspheres can be avoided.
2) PBA can be prepared with good reproducibility by an easy-to-handle method. No problems are foreseen on the scale-up of the manufacturing method.
3) Since the particles are composed of pure bioactive compounds and a very small amount of stabilizer, the drug loading ability of the drug particles is great.

4) Release of pharmaceuticals or other biologically active compounds from the formulation can be controlled by selection of the amphiphilic compound used to stabilize the particles.
5) The preparation of PBA can avoid the use of toxicologically active additives.
6) For example, freeze-dried PBA dispersion can produce a moisture-free storage system with increased stability.
7) The surface properties of PBA are modified by the selection of amphiphilic compounds used as stabilizers and also by attaching so-called homing devices, eg monoclonal antibodies or carbohydrate moieties, for targeting drugs. can do. The surface modification modifies bioavailability and biodistribution with respect to the degree and rate of absorption, circulation time, site of action and excretion mode of the bioactive substance. Modification of surface properties provides an opportunity to avoid or at least reduce uptake by intravenously administered particles of RES cells.

8) Since particles with a particle size below 100 nm-200 nm can be prepared, they have an opportunity to overflow from the endothelium wall window. This allows the drug to be targeted to an extravascular location, such as the bone marrow.
9) Since the particle size is reduced to a nanometer particle size range that cannot generally be achieved by grinding or grinding, the specific surface area of the particles increases tremendously. Since the oral bioavailability of a drug or other bioactive substance is related to the specific surface area by the dissolution rate of the substance in the gastrointestinal tract, submicron sized particles enhance the bioavailability of drugs that are sparingly soluble in GIT .
10) Hydrophobic substances can be formulated as PBA having a hydrophilic surface. A hydrophilic surface, for example in GIT, provides good wettability of the particles and facilitates dissolution of the compound. That is, bioavailability can be increased.
11) PBA manufacturing methods include only inexpensive and easy-to-handle methods and give products that are safe to handle. Due to the presence of the particles in the liquid dispersion, there is no danger of dust explosion, cross-contamination or inhalation of bioactive substances as encountered during the production of very fine powders.

The invention relates to suspensions of micron and submicron particles of solid biodegradable lipids (solid lipid particles, SLP), suspensions of fusible bioactive agent particles (PBA), lyophilizates thereof and The present invention relates to a manufacturing method thereof.
Solid lipid particles (SLP) have a predominantly non-equiaxial shape, which is similar to that of β-polymorphic (eg β ′, β ₁ , β ₂ ) lipid matrix or triglyceride β-crystals The result is that it exists in the polymorphic state and not in the amorphous or α-crystal-like state. SLP is intended for parenteral administration of poorly water-soluble substances such as drugs or other biologically active substances, but also for oral, nasal, pulmonary, rectal, direct skin and buccal administration Can be used as a carrier system. However, the application of SLP is not limited to pharmaceutical administration to humans or animals. SLP can also be used in cosmetics, food and agricultural products. SLP is a new lipid structure with properties that overcome many of the problems associated with previously described carrier systems.

  The matrix of SLP is composed of a biocompatible (biocompatible) hydrophobic material that is solid at room temperature and has a melting point in the range of about 30-120 ° C. Preferred matrix components are solid lipids (fats) such as mono-, di- and triglycerides of long chain fatty acids; hydrogenated vegetable oils; fatty acids and their esters; fatty alcohols and their esters and ethers; natural or synthetic waxes such as beeswax and Carnauba wax; wax alcohols and their esters, sterols such as cholesterol and its esters, solid paraffin, and mixtures thereof. The carrier material must be compatible with the agent to be incorporated.

  Lipids are known to exhibit significant polymorphism. This can be defined as the ability to reveal different unit cell structures in the crystal that come from various molecular conformations and molecular packings. Depending on the conditions, for example, glycerides are classified according to Larsson's classification (K. Larsson, 1966, Acta Chem. Scand. 20, 2255-2260), alpha (α), beta prime (β ′) and beta (β ) And can be crystallized in three different polymorphic forms. These polymorphic modifications characterized by specific carbon chain loading can differ significantly in their properties such as solubility, melting point and thermal stability. The transformation occurs as α → β ′ → β, and the transition is monotropic. Depending on thermal conditions, the β-form is the most thermodynamically stable polymorph, whereas α is the least stable and to some extent more rapidly the more stable polymorphs β ′ and β To metamorphosis. This transformation is accompanied by changes in physicochemical properties.

  In the aforementioned suspension of SLP, the lipid matrix exists mainly in a stable polymorphic modification. Upon cooling, a dispersed melt metastable polymorph, such as the α-form, can immediately form, but a stable polymorph is formed within hours or days after the dispersion is prepared.

  SLP suspensions can be stabilized with amphiphilic compounds such as ionic and non-ionic surfactants. Suitable stabilizers include, but are not limited to, the following: natural and synthetic phospholipids, their hydrogenated derivatives and mixtures thereof, sphingolipids and glycosphingolipids; physiological bile acids Salts such as sodium cholate, sodium dehydrocholate, sodium deoxycholate, sodium glycocholate and sodium taurocholate; saturated and unsaturated fatty acids or fatty alcohols; ethoxylated fatty acids or fatty alcohols and their esters and ethers; Polyether alcohols such as tyloxapol; esters and ethers of sugars or sugar alcohols with fatty acids or fatty alcohols; acetylated or ethoxylated mono- and diglycerides; examples of synthetic biodegradable polymers If a block copolymer of polyoxyethylene and polyoxypropylene oxide; ethoxylated sorbitan ester or sorbitan ether; amino, polypeptides and proteins such as gelatin and albumin; or two or more combinations of the foregoing.

  The aqueous phase in which the SLP is dispersed is a water-soluble or dispersible stabilizer; isotonic agents such as glycerol or xylitol; cryoprotectants such as sucrose, glucose, trehalose, etc .; electrolytes; buffers; Sodium acid, sodium pyrophosphate or sodium dodecyl sulfate; can contain preservatives.

Depending on the nature of the stabilizer used, other colloidal structures such as micelles and vesicles may coexist in the suspension of SLPs.
Substances that are particularly suitable for capture in SLP are poorly water-soluble, exhibit low bioavailability, poor absorption from the intestine, and / or more rapidly by chemical or enzymatic processes in the biological environment Pharmaceuticals or other biologically active compounds that will be degraded to low levels, and low specific active substances that are extremely toxic at non-target sites. If it is desired to incorporate a relatively water-soluble compound into the SLP, it is necessary to reduce the water solubility of the compound, for example, a water-insoluble derivative of the compound, such as an acid or base, complex, or parent. It can be achieved using oily precursors and the like.

  Drugs or bioactive agents that are particularly suitable for incorporation into SLPs include antibiotics such as fosfomycin, fosmidomycin and rifapentin; antihypertensive drugs such as minoxidil, dihydroergotoxin ( dihydro-ergotoxine and endralazine; antihypertensive drugs such as dihydroergotamine; systemic antifungal drugs such as ketoconazole and keroseconazole and griseofulvin; anti-inflammatory drugs such as indomethacin and diclofenac , Ibuprofen, ketoprofen and pirprofen; antiviral drugs such as acyclovir, vidarabine and immunoglobulin; ACE inhibitors such as captopril And enalapril; beta blockers such as proprano-lol, atenolol, metoprolol, pindolol, oxprenolol and labetalol; bronchodilators such as ipratropium bromide (Ipratropiumbromide) and sobrerol; calcium antagonists such as diltiazem, flunarizin, verapamil, nifedipine, nimodipin and nitrendipin; cardiac glycosides such as digitoxin digitoxin, digoxin, methyldigoxin and acetyl-digoxin; cephalosporins such as ceftizoxim, cefalexi n), cefalotin and cefotaxim; cytostatic drugs such as chlormethin, cyclophosphamid, chlorambucil, cytarabin, vincristin, mitomycin C ), Doxorubicin, bleomycin, cisplatin, taxol, penclomedine and estramustin; hypnotics such as flurazepam, nitrazepam and lorazepam Psychotropic drugs such as oxazepam, diazepam and bram-azepam; steroid hormones such as cortisone, hydrocortisone, prednisone, pudnisone, dexamethasone, proge Teron, pregnanolone, testosterone and testosterone undecanoate; vasodilators such as molsidomin, hydralazin and dihydralazin; cerebral vasodilators such as dihydroergotoxin, ciclonicat and vincamin ); Lipophilic vitamins such as vitamins A, D, E, K and their derivatives.

  Bioactive substances can be located in the core of SLP where they are dissolved, solubilized or dispersed in the matrix and / or in the stabilizer layer surrounding the particle matrix and / or adsorb to the surface of SLPs Can do. The bioactive material may be dissolved or in a crystalline or amorphous state, or a mixture of these crystallographic states.

SLPs are somewhat similar to the preparation of lipid-in-water (oil) emulsions, but can be prepared by an emulsification method characterized primarily by their fundamental differences as outlined below. The method is described as follows:
(1) Melt solid lipid or lipid mixture.
(2) Add stabilizers to the lipid and dispersion medium, or to the dispersion medium only, depending on their physicochemical properties. The choice of stabilizer and mixing regime is not comparable to that applied to lipid-in-water (oil) emulsions, as will be apparent from the examples below. Stabilizers can also be added or exchanged after homogenization, for example by polymer adsorption or by dialysis of water-soluble surfactants.
(3) Drugs or other bioactive substances to be incorporated into the SLP may be melted together with lipids if the physicochemical properties of the substance permit, or dissolved and solubilized in the lipid melt prior to homogenization Or it may be dispersed.

(4) The dispersion medium is heated to the temperature of the melt before mixing and may contain, for example, stabilizers, isotonic agents, buffer substances, cryoprotectants and / or preservatives.
(5) The melted lipid compound is emulsified in the dispersion medium, preferably by high pressure homogenization, but emulsification can also be achieved by sonication, high speed stirring, vortexing and vigorous hand shaking. The particle size of the SLPs is determined by the homogenization method.
The basic differences for lipid-in-water emulsions other than the choice of stabilizer and mixing regime relate to the following steps:
(6) After homogenization, the dispersion can be sterilized by standard methods such as autoclaving, or filtration through a 0.2 μm sterilizing filter if the particles are small enough not to be retained by the filter. These steps must be performed before the system is cooled below the recrystallization temperature. Furthermore, contamination that can result in heterogeneous nucleation should be avoided. Therefore, it is better to remove the particulate contaminants from the dispersion by cooling before cooling to a temperature lower than the recrystallization temperature. The pore size of the filter should be chosen large enough so as not to retain lipid particles.

(7) When the dispersion is allowed to cool at room temperature, SLP is formed by recrystallization of the dispersed lipid. During cooling, the dispersion may be stirred, for example with a magnetic stirrer.
(8) In the next step, the amount of dispersion medium can be reduced, for example by evaporation, or the dispersion medium can be removed by standard methods, for example filtration, ultrafiltration or lyophilization, so that it can be reconstituted before use. A water-free storage system is obtained which can be constructed. The lyophilized powder can also be processed into other pharmaceutical, cosmetic, food or agricultural dosage forms such as powders, tablets, capsules and the like.

  As demonstrated by FIG. 1, which shows a transmission electron micrograph of a frozen cleaved specimen of the SLP of Example 1, SLPs are typically non-equiaxial solid particles. Non- equiaxed particle shape results from the lipid matrix crystallizing as a β-polymorphic form. Solidification of amorphous fat or crystallization of unstable α-polymorph generally reveals spherical particles. The presence of stable β-form could be detected by differential scanning calorimetry (FIG. 2) and synchrotron radiation wide angle X-ray diffraction (FIG. 3).

The particle size of SLP depends on the type and amount of emulsifier and the method and conditions of emulsification (see below).
Re-solidification of the molten lipid prior to homogenization should be avoided. This is because, if the particles are solid during this process, the diameter reduction by homogenization is substantially prevented. In order to ensure that the molten lipid does not solidify before homogenization, i.e. before small particles can be formed, the two phases are mixed after heating the dispersion medium to approximately the same temperature as the melt. Therefore, the melt is not cooled by the addition of the dispersion medium.

  SLPs in the nanometer size range can be obtained by high pressure homogenization. The particles exhibit a relatively narrow particle size distribution with an average particle size of about 50-300 nm as measured by photon correlation spectroscopy (PCS). SLP dispersions are stable on storage for over 18 months. That is, long-term stability is similar to that of submicron o / w emulsions used for parenteral nutrition purposes. Patents such as Liposphere (A. Domb et al, International Application No. PCT / US90 / 06519, November 8, 1990) and Lipid Nanopellets (P. Speiser, European Patent No. 0167825, issued on August 8, 1990) Long-term stability data for other solid lipid-based carrier dispersions described in the literature could not be found. Domb describes phospholipid-stabilized tristearate lipospheres that are stable for 7 days as "exceptionally stable".

  To stabilize the SLP suspension, ensure that the amount of high mobility stabilizer in the dispersion medium is sufficient to stabilize the newly generated surface during recrystallization after emulsification. It was found that a high mobility stabilizer must be present in the dispersion medium (see below). It has been found that bile salts are very efficient in this respect when combined with non-electrolytic compounds such as glycerol, particularly at concentrations used to achieve blood isotonicity. SLPs stabilized by phospholipids alone or in combination with non-electrolyte compounds such as glycerol at concentrations used to achieve blood isotonicity are shown in the transmission electron micrograph of FIG. Tends to form a semi-solid, ointment-like gel, whereas addition of sodium glycocholate to the aqueous phase prevents this gel formation (B. Siekmann and K. Westesen, 1992, Pharm. Pharmacol. Lett 1, 123-126).

A phospholipid: bile salt molar ratio of 2: 1 to 4: 1 has been found to be extremely effective with respect to initial stabilization during homogenization and also with respect to long-term storage stability of SLP dispersions. These phospholipid / bile acidifications are higher than the mixed micelle formation ratio and correspond to the swollen lamellar phase of the mixed lecithin / bile salt layer in the lecithin / bile salt / water ternary phase diagram. Thus, the data show that stabilization is very effective when the bile salt is not bound to mixed micelles, and bile salt molecules are inserted into the phospholipid layer on the particle surface during SLP stabilization. Suggests that
SLPs can also be sterically stabilized by nonionic surfactants. However, steric stabilization of SLP requires a relatively large amount of surfactant with a lipid / surfactant ratio of 1: 1 or less. In general, it can be seen that the stability of SLPs decreases with increasing lipid / surfactant ratio.

  The amount of emulsifier required for surface stabilization of the dispersed particles is higher than in conventional lipid emulsions such as those used for parenteral nutrition. This effect can be attributed to molten lipid crystallization after homogenization. Lipids are typically recrystallized or present (on storage) as non-equiaxial particles rather than ideal spheres, resulting in a significant increase in surface area compared to droplets of emulsified molten lipids or conventional lipid emulsions, respectively. To do. Additional surfaces that regenerate during dispersed lipid recrystallization or polymorphic transitions need to be stabilized immediately after formation to avoid particle aggregation. Therefore, the preparation of a stable SLP dispersion requires the presence of a stabilizer reservoir after emulsification.

The choice of stabilizer cannot be inferred from the composition of oil-in-water emulsions and the stabilization mechanism, and depends on the presence of highly mobile stabilizers due to the mechanism of formation of non-equiaxial particles. In colloid and surface science, “high mobility” generally refers to free diffusion at a high diffusion rate in a dispersion medium. With regard to the stabilization of colloidal solid lipid particles, in order to exert a stabilizing action at the lipid / water interface to prevent particle aggregation, the diffusion rate is increased before particle aggregation can occur (especially during lipid recrystallization). B) should be high enough to reach the nascent particle surface. A sufficiently high diffusion rate is typically observed for materials that do not form a phase separation in the dispersion medium according to the phase rule set by Gibbs. High mobility stabilizers may be ionic or non-ionic. Typically, these stabilizers are molecularly dissolved in the dispersion medium and / or form micelles. Micelles are known to be highly dynamic structures characterized by fast molecular exchange between micelle aggregates and dispersion media. Monomers in the dispersion medium are readily available for surface stabilization. In contrast, stabilizers that tend to form a phase separation in the dispersion medium are not sufficiently mobile to stabilize the nascent surface before particle agglomeration can occur. Therefore, these stabilizers are not suitable as sole stabilizers for SLP dispersions. Phospholipids are an example of a stabilizer that forms a phase separation in a dispersion medium. It is well known that phospholipids form a closed layered structure in an aqueous medium, so-called vesicles, and that the exchange rate of phospholipid molecules between the vesicle and the aqueous phase is very small compared to that of micelles. . Thus, phospholipid molecules are attached to the vesicle structure and are not readily available to stabilize the nascent surface during recrystallization of lipid particles. Thus, as is apparent from Examples 5 and 13, even though phospholipids alone are suitable as effective stabilizers for lipid emulsions, they cannot efficiently stabilize SLP suspensions. Indeed, preparing SLPs with standard compositions of lipid emulsions, such as 10% fat and 1.2% phospholipids, results in unstable SLP dispersions. Even general and high concentrations of phospholipids, for example 20% or 60% lecithin, for the fatty phase, as demonstrated in Example 5, are not sufficient to stabilize the SLP dispersion.

Although described in patent literature (eg Domb et al, US Patent Application No. 435,546 filed November 13, 1989 and International Application No. PCT / US90 / 06519 filed November 8, 1990), A fine suspension of lipids is not equivalent to a submicron lipid emulsion in that its internal phase is not replaced by liquid but only by solid fat. The physicochemical properties of lipid suspensions such as SLP are substantially different from those of lipid emulsions. Because of these differences, lipid suspensions cannot be prepared as well as lipid emulsions. One fundamental difference between SLPs is that their particle surface area is much larger, as already outlined, so SLPs require higher amounts of surfactant, and the surfactant further In order to immediately stabilize the nascent surface when the molten lipid is recrystallized or transformed into a stable β-polymorph, it must be highly mobile. The second basic difference is that once the SLP is formed by recrystallization of the molten lipid, there is no excess mobile stabilizer in the aqueous phase that is not adsorbed on the particle surface where it is immobilized. When the small particle melting is renewed, the dispersion becomes unstable. Another requirement for the physicochemical stability of SLPs as opposed to oil-in-water emulsions is the absence of particulate impurities that can promote heterogeneous nucleation. Therefore, it is better to remove the particulate contaminants from the dispersion by filtration before cooling to a temperature lower than the recrystallization temperature. Furthermore, it has been found that non-electrolytic compounds used to achieve blood isotonicity, such as glycerol, promote the stability of SLP dispersions.
The invention also relates to a suspension of bioactive agent particles (PBA). Poorly water-soluble substances that can be melted in the temperature range of about 30 to 120 ° C., such as pharmaceuticals, insecticides, antifungal agents, pesticides, herbicides, fertilizers, nutrients, cosmetics, etc. Can be formulated as PBA. The PBA matrix is composed of the bioactive agent itself.

  PBA provides a new type of delivery system and can be characterized as primarily submicron and / or micron particles of bioactive agent suspended in an aqueous medium that is stabilized by an amphiphilic compound. PBA has modified surface properties that can be controlled by the choice of amphiphile and / or a matrix-constituting compound with a reduced particle size compared to particulate material. These properties modify the biodistribution and bioavailability of the formulated drug or other bioactive substance, which means the degree and rate of dissolution and absorption of the drug or other bioactive substance. Means that the circulation time, the site of action and the mode of excretion are modified. The physicochemical properties of PBA greatly depend on the characteristics of the bioactive agent that is a constituent of PBA, the type and amount of the stabilizer, and the emulsification method. PBA suspensions and lyophilizates are used for oral, nasal, pulmonary, rectal, transdermal, buccal administration of poorly water-soluble drugs or other bioactive compounds, and, depending on the particle size, parenteral It can also be used for administration. Furthermore, PBA can also be used in cosmetics, foods and agricultural products, in particular to make up poorly water-soluble herbicides and pesticides.

The matrix of PBA preferably has a melting point below 100 ° C, or it can be lowered to a temperature below 100 ° C by the addition of certain adjuvants, by practically insoluble or poorly water soluble agents. Composed. Substances that are particularly suitable for composition as PBA are drugs or other bioactive substances that are poorly water soluble, exhibit low bioavailability, and / or have poor absorption from the intestine. Examples of such materials include, but are not limited to:
Anesthetics and narcotics such as butanilicaine, fomocaine, isobutambene, lidocaine, risocaine, prilocaine, pleudocaine, pseudococaine, tetracaine, tetracaine trimecaine, tropacocaine and etomidate; anticholinergics such as methixen and profenamine; antidepressants, psychostimulants and neuroleptics such as alimenazine, binedaline , Perazine, chlorpromazine, fenpentadiol, fenanisol, fluanisol, mebenazine, methylphenidate, thioridazine , Toloxaton and trimipramine; antiepileptic drugs such as dimethadion and nicetamide; antifungal drugs such as butoconazole, chlorphenesin, etisazole, exalamid , Pecilocine and miconazole; anti-inflammatory drugs such as butibufen and ibuprofen; bronchodilators such as bamifylline; cardiovascular drugs such as alprenolol, butobendine, chloridazole , Hexobendine, nicofibrate, penbutolol, pirmenol, prenylamine, procaine amide, propatinyl nitrate Propatylnitrate, suloctidil, trilipolol, xibendol and biquidile; cytostatic drugs such as asperline, chlorambucil, chlornaphazine, mitotane, est Ramustine, taxol, penchromedin and trofosfamide; hyperemic agents such as capsaicin and methyl nicotinate; lipid lowering agents such as nicoclonate, oxprenolol, pirifibrate, simfib Simfibrate and thiadenol; analgesics such as aminopromazine, caronerine, difamerine, fencarbamide, tiropramide and moxibustion Testosterone derivatives such as testosterone enanthate and testosterone- (4-methylpentanoate); tranquilizers such as azaperone and buramate; virus quiescent drugs such as arildon; vitamin A derivatives; Retinol, retinol acetate and retinol palmitate; vitamin E derivatives such as tocopherol acetate, tocopherol succinate and tocopherol nicotinate; menadione; cholecalciferol; insecticides, pesticides and herbicides such as acephate ), Cyfluthrin, azinphosphomethyl, cypermethrine, substituted phenylthiophosphate, fenclop hos), permethrine, piperonal, tetramethrine and trifluraline.

  As with SLP, a suspension of PBA can be stabilized with an amphiphilic compound. In principle, the same ionic and nonionic surfactants that can be used to stabilize SLPs are also suitable for the preparation of PBA suspensions. The choice of stabilizer depends on the physicochemical properties of both the bioactive material and the dispersion medium and the desired surface properties of the particles.

  The aqueous phase in which the PBA is dispersed is a water-soluble (or dispersible) stabilizer: isotonic agents such as glycerol or xylitol; cryoprotectants such as sucrose, glucose, trehalose, etc .; electrolytes; buffers; aggregation inhibitors such as sodium citrate Sodium pyrophosphate or sodium dodecyl sulfate; should contain preservatives. Although water is a preferred dispersion medium, the invention is not limited to aqueous dispersions, and any other pharmacologically acceptable liquid such as ethanol, propylene glycol and methyl-isobutyl-ketone or their Can be spread into the mixture.

Depending on the properties of the stabilizer used, other colloidal structures such as micelles and vesicles may coexist in the PBA suspension.
The suspension of PBA is typically prepared by the same emulsification method as in SLP. The molten pharmaceutical or bioactive substance or mixture of such compounds is emulsified, preferably by high pressure homogenization, in a pharmacologically acceptable liquid that is immiscible with the melt. Emulsification is also possible by sonication, high speed stirring, vortexing and vigorous hand shaking. The liquid is heated to the melt temperature prior to mixing and may contain, for example, isotonic agents, buffer substances, cryoprotectants and / or preservatives.

Stabilized amphiphilic compounds are added to the melt and liquid, or to the liquid only, depending on their physicochemical properties. Stabilizers can also be added or exchanged after homogenization, for example by polymer adsorption or by dialysis.
The PBAs produced by the above method can be divided into two distinct groups.
The characteristics of the first group of PBAs are that they are water insoluble at the emulsion preparation temperature and are not solubilized by excess stabilizer or form micelles themselves, and the PBA particle size is room temperature. It remains in the same state before and after cooling.
A characteristic of the second group of PBAs is that they are partially water soluble at the emulsion preparation temperature and / or can form mixed micelles with excess stabilizer and / or form micelles on their own. As a result, substances from small to large particles, for example by crystal growth and / or precipitation of dissolved bioactive agents from supersaturated solutions and / or by methods such as micelles and / or Ostwald ripening, for example By transportation, the particle size increases after cooling to room temperature.

In the next step, the liquid phase can be removed, for example by lyophilization, thereby producing a reconstitutable powder that can also be processed into other pharmaceutical formulations.
PBAs are finely dispersed particles composed of a matrix of bioactive material surrounded by one or more layers rich in surfactants. Its particle size and size distribution, and particle shape and surfactant coating depend on the nature and amount of the matrix-forming bioactive agent and stabilizer, the bioactive agent to amphiphile ratio, and the emulsification method.

Example 1 Method for Preparing Tripalmitate SLP In a thermostatic glass bottle, 4.0 g of tripalmitate (tripalmitin, 99% purity, Fluka) is heated to 75 ° C. to melt the lipid. 0.48 g of soy lecithin (Lipoid S 100, Lipoid KG) is dispersed in the lipid melt by probe sonication (MSE Soniprep 150) until the dispersion is optically clear. 0.16 g sodium glycocholate (glycocholic acid, sodium salt 99%, Sigma) and 4 mg thiomersal (Synopharm) are dissolved in 36 ml double distilled water. The aqueous phase is heated to 75 ° C. and added to the lipid melt. A coarse dispersion is produced by probe sonication for about 2 minutes. The crude dispersion is transferred to a constant temperature high pressure homogenizer (APV Gaulin Micron Lab 40) and passed 5 times through the homogenizer at a pressure of 500 bar. Homogenization with this device is performed by extruding through a small ring orifice. The homogenized dispersion is allowed to cool at room temperature. The trace amount of visible fat particles exhibited by the dispersion is separated from the dispersion by filtering it through a 0.45 μm sterile filter.

The importance of mixing highly mobile surfactants such as bile salts with respect to particle size distribution and SLP dispersion stability is demonstrated below, for example in Examples 2, 5-7 and 13-15.
The average particle size after preparation (by number) of tripalmitate SLP measured by photon correlation spectroscopy (PCS, Malvern Zetasizer 3) is 205 nm. Even after 15 months of storage, the particles show no visible signs of agglomeration, creaming, settling or phase separation. PCS polygon measurement (Malvern Zetasizer 3, detected at five different angles, ie 50, 70, 90, 110 and 130 °) shows a monomodal (by number) size distribution with a peak at 250 nm ( FIG. 5).
At temperatures below the melting point of the lipid matrix, it is primarily non-equal axis particles, as demonstrated in FIG. 1, which is a transmission electron micrograph of a frozen cleaved specimen of the tripalmitate SLP of Example 1. Prior to preparation of the specimen, the sample is stored at room temperature for 5 months. The sample is frozen and cleaved at 173 K with a freezing cleaving apparatus BAF 400 (Balzers AG, CH-Liechtenstein). Fast freezing is performed by slushing in molten propane. Samples are shadowed using platinum / carbon (layer thickness 2 nm) at 45 ° and pure carbon at 90 ° to prepare replicas. The replica is cleaned with a 1: 1 (v / v) chloroform / ethanol mixture. The replica on the uncoated grid is observed using an electron microscope EM 300 (Philips, D-Kassel).

  In non-equal axis tripalmitate particles, glyceride exists as a stable β-crystal polymorph as shown by thermal analysis studies. FIG. 2 shows the differential scanning calorimetry (DSC) thermogram of the SLP of Example 1 and of pure tripalmitate. The samples are accurately weighed into a standard aluminum pan. Thermograms are recorded from 20 ° C to 90 ° C at a scan rate of 10 ° C / min using a Perkin Elmer calorimeter DSC-7. The detected transition peak corresponds to the melting of the tripalmitate β-crystal. The melting point of tripalmitate SLP moves to a lower temperature than that of pure tripalmitate due to the presence of phospholipids and the small crystallite size.

Example 2 : Method for preparing isotonic tripalmitate SLP
Melt 7.0 g of tripalmitate (tripalmitin, Fluka) in a glass bottle thermostatized at 75 ° C. 840 mg soy lecithin (Lipoid S 100, Lipoid KG) is dispersed in the tripalmitate melt as described in Example 1. An aqueous phase containing 1.575 g glycerol, 280 mg sodium glycocholate and 4 mg thiomersal is heated to 75 ° C. and added to the lipid melt to a weight of 70 g. A coarse dispersion is produced by sonication for about 2 minutes. The crude dispersion is transferred to a constant temperature high pressure homogenizer (APVGaulin Micron Lab 40) and passed 10 times through the homogenizer at a pressure of 800 bar. The homogenized dispersion is allowed to cool at room temperature.
The average particle size according to the number of isotonic tripalmitate SLPs measured by PCS is 125.9 nm after preparation and 116.2 nm after storage for 50 days, ie practically no particle growth. Small deviations in the numerical values fall within the accuracy range of the particle size measurement method. In FIG. 6, the PCS particle size distribution is compared to that of Intralipid ^™ , a commercially available lipid emulsion for parenteral nutrition. Intralipid ^™ consists of 10% soybean oil, 1.2% fractionated phospholipids and 2.25% glycerol dispersed in water for injection. It can be seen that the particle size distribution of the tripalmitate SLP of Example 2 is significantly smaller and narrower than that of Intralipid ^™ . In contrast to Example 1, the addition of glycerol produces a significant difference in particle size distribution. The SLP dispersion of Example 1 contains a small amount of visible suspension particles after cooling from the hot emulsion to room temperature, whereas the dispersion of Example 2 is macroscopically visible suspension. No liquid particles are seen.
Investigation by synchrotron radiation wide-angle X-ray diffraction (FIG. 3) and differential scanning calorimetry shows that tripalmitate in SLP exists as a stable β-polymorph at room temperature.

Example 3 : Preparation of hard fat SLP by microfluidization
3.0 g of hard fat (Witepsol ^™ W35, Huels AG) is melted at 75 ° C. in a thermostatic glass bottle. 1.8 g soy lecithin (Phospholipon 100, Natterman) is dispersed in the tripalmitate melt as described in Example 1. An aqueous phase containing 375 mg sodium glycocholate, 2.25 g glycerol and 10 mg thiomersal is heated to 75 ° C. and added to the lipid melt to a weight of 100 g. A coarse dispersion is produced by ultra-turrax vortexing for about 2 minutes. The coarse dispersion is transferred to a microfluidizer (Microfluidics Microfluidizer M-110T), which is a high-pressure homogenizer based on the jet flow method, immersed in a constant temperature water bath (70 ° C.). The dispersion is circulated through the microfluidizer for 10 minutes and allowed to cool at room temperature.
The average particle size of the prepared hard fat SLP as measured by PCS is 45.9 nm.
In order to monitor the time course of homogenization, a sample for particle size measurement is taken every minute during homogenization. FIG. 7 shows the average particle size versus homogenization time. The average particle size decreases with time and does not change at the end of homogenization.

Example 4 : Long-term stability of hard fat SLPs prepared by microfluidization The stability of hard fat SLPs is monitored over a year. During this period, the sample is stored in the refrigerator at about + 4 ° C. The particle size distribution of the sample is measured by PCS at regular time intervals. FIG. 8 demonstrates that the average particle size of hard fat SLP is practically constant over a one year monitoring period.

Example 5 : Preparation of unstable SLP dispersion 4.0 g of tripalmitate (Dynasan 116, Huels AG) is heated to 75 ° C in a thermostatic glass bottle to melt the lipid. 0.48 g of soy lecithin (Lipoid S 100, Lipoid KG) is dispersed in the lipid melt by probe sonication (MSESoniprep 150) until the dispersion is optically clear. 4 mg thiomersal and 0.9 g glycerol are dissolved in 35.6 ml double distilled water. The aqueous phase is heated to 75 ° C. and added to the lipid melt. A coarse dispersion is produced by probe sonication for about 2 minutes. The crude dispersion is transferred to an isothermal high pressure homogenizer (APV Gaulin Micron Lab 40) and passed 5 times through the homogenizer at a pressure of 500 bar. The homogenized dispersion is allowed to cool at room temperature. When stored, the SLP dispersion becomes a milky, semi-solid, cartilaginous gel.
3.0 g of hard fat (Witepsol ^™ W35, Huels AG) is melted at 75 ° C. in a thermostatic glass bottle. 1.8 g soy lecithin (Phospholipon 100, Natterman) is dispersed in the tripalmitate melt as described in Example 1. An aqueous phase containing 10 mg thiomersal is heated to 75 ° C. and added to the lipid melt to a weight of 100 g. A coarse dispersion is produced by ultra-turrax vortexing for about 2 minutes. The coarse dispersion is transferred to a microfluidizer (Microfluidics Microfluidizer M-110T) immersed in a constant temperature water bath (70 ° C.). The dispersion is circulated through the microfluidizer for 10 minutes and allowed to cool at room temperature. When stored, the SLP dispersion becomes a cloudy semi-solid, ointment-like gel. A transmission electron micrograph of this gel is shown in FIG.
In a thermostatic bottle, 4.0 g of tripalmitate (Dynasan 116, Huels AG) is heated to 80 ° C. to melt the lipid. 0.8 g of soy lecithin mixture (Lipoid S 75, Lipoid KG) is dispersed in the lipid melt by probe sonication (MSE Soniprep 150) until the dispersion is optically clear. 4 mg thiomersal is dissolved in 35.6 ml double distilled water. The aqueous phase is heated to 80 ° C. and added to the lipid melt. A coarse dispersion is produced by probe sonication for about 2 minutes. The crude dispersion is transferred to a constant temperature and high pressure homogenizer (APV Gaulin Micron Lab 40) and passed 5 times through the homogenizer at a pressure of 800 bar. The homogenized dispersion is filled into a glass bottle and allowed to cool at room temperature. When the dispersion is cooled to room temperature, it forms fat aggregates that appear semi-solid on the walls of the glass bottle. This dispersion gels when a shear force is applied, for example by passing it through a hypodermic syringe.
The use of phospholipids alone as a stabilizer, such as found in commercially available parenteral oil-in-water emulsions, clearly does not result in a stable system in the case of SLP suspensions. Even using phospholipids such as Lipoid S 75, which induce a fairly high negative net charge, cannot provide sufficient stabilization. As further outlined in Examples 6, 7, and 13, electrostatic repulsion alone cannot be the basic stabilization mechanism for SLP.

Example 6: Preparation of tripalmitate SLP sterically stabilized with tyloxapol A series of tripalmitate SLP dispersions stabilized with tyloxapol (Eastman Kodak) is prepared with varying lipid / surfactant ratio. These SLP dispersions are produced according to the following procedure:
Dissolve tyloxapol in heated double distilled water while keeping the temperature below the cloud point of tyloxapol (about 90-95 ° C.). Tyloxapol solution at a temperature of 80 ° C. is added to the molten tripalmitate or tripalmitate / lecithin dispersion at the same temperature, respectively. A crude emulsion is produced by probe sonication for about 2 minutes. The crude emulsion is then passed 5 times through a high-pressure homogenizer at a pressure of 1200 bar. The homogenized dispersion is allowed to cool at room temperature. All dispersions contain 2.5% glycerol and 0.01% thiomersal.
Table 1 shows the composition of the prepared SLP dispersions and the average particle size (by number) after their preparation as measured by PCS. The asterisk (*) in the particle size column indicates that the dispersions exhibit a bimodal particle size distribution with a particle size that is considerably larger than indicated by the average particle size. It can be seen that a sterically stabilized SLP dispersion requires a large amount of surfactant to obtain a uniform particle size SLP. In the case of SLP stabilized by tyloxapol and phospholipid, the surfactant ratio needs to be optimized. In this series, it was found that a tyloxapol / lecithin ratio of at least 1: 1 gave SLPs of uniform particle size. As the ratio increases, the average particle size decreases. As in Examples 1 to 3, to obtain a stable dispersion, it is necessary to add a highly mobile surfactant capable of forming micelles. A large amount of surfactant is required to create a surfactant reservoir in the dispersion medium that can provide sufficient surfactant molecules when the molten lipid recrystallizes and forms non-equal axis particles with a large specific surface area. Become.

Example 7 : Preparation of tripalmitate SLP sterically stabilized with poloxamer
1.2 g soy lecithin (Lipoid S 100, Lipoid KG) is dispersed in 4.0 g molten tripalmitate (Dynasan 116, Huels AG) by probe sonication at a temperature of 80 ° C. 1.8 g poloxamer (Pluronic ^™ F68, BASF), 0.9 g glycerol and 4 mg thiomersal are dissolved in 32.1 g double-distilled water heated to 80 ° C. The heated solution is added to the lipid melt and a crude dispersion is prepared by probe sonication for 2 minutes. The crude dispersion is transferred to a constant temperature, high pressure homogenizer (APV Gaulin Micron Lab 40) and passed 5 times through the homogenizer at a pressure of 1200 bar. The homogenized dispersion is allowed to cool at room temperature.
This poloxamer-stabilized SLP exhibits a single-numbered particle size distribution with average particle size (by number) after preparation as measured by 77.9 nm PCS. Due to the presence of an excess of highly mobile surfactant in the aqueous phase, this system is stabilized during recrystallization of the molten lipid and gelled as seen with systems stabilized solely by phospholipids. Does not happen.

Example 8 : Effect of homogenization pressure on average particle size of SLP SLPs having the following composition are prepared at various homogenization pressures. These SLP dispersions are 3% tripalmitate (Dynasan 116, Huels AG), 1.5% tyloxapol, 1% soy lecithin (Lipoid S 100, Lipoid KG), 0.01% thiomersal and 100% total. Consists of double distilled water (% is weight). The lecithin is dispersed in molten tripalmitate (80 ° C.) by probe sonication until the dispersion is optically transparent. Tyloxapol is dissolved in warm water (80 ° C) containing thiomersal. The SLP dispersion is prepared as described in Example 6.
FIG. 9 shows the effect of homogenization pressure on the average particle size of SLP. As pressure increases, the particle size decreases and the particle size distribution narrows.

Example 9 : Effect of pass through homogenization on mean particle size of SLP 3% tripalmitate (Dynasan 116, Huels AG), 1.5% tyloxapol, 1% soy lecithin (Lipoid S 100, Lipoid KG) A tripalmitate SLP composed of 0.01% thiomersal and double distilled water (% by weight) with a total amount of 100% is prepared as described in Example 6 at a pressure of 800 bar. Samples for particle size measurement are taken after preparation of the crude emulsion and after each pass through a homogenizer.
FIG. 10 shows the influence of the number of passes on homogenization on the average particle size of SLP, and the average particle size decreases as the number of passes increases.

Example 10 : Preparation of SLP by probe sonication-Effect of sonication time on average particle size of SLP
Melt 1.20 g of tripalmitate in a glass bottle for sonication at 80 ° C. 0.40 g of soy lecithin (LipoidS 100) is dispersed in the lipid melt by probe sonication until the dispersion is optically clear. 0.60 g tyloxapol and 4 mg thiomersal are dissolved in double distilled water heated to 80 ° C. The aqueous phase is added to the lipid melt and an SLP dispersion is prepared by probe sonication at 80 ° C. The sonication is operated at 50% of its maximum power. At regular time intervals (1, 5, 10 and 15 minutes), samples are taken from the dispersion for particle size measurement. After 30 minutes, probe sonication is stopped and the dispersion is allowed to cool at room temperature.
The influence of the sonication time on the average particle size of SLP is shown in FIG. As the sonication time increases, the average particle size decreases and the particle size distribution narrows.

Example 11 : Preparation of SLP by stirring An SLP dispersion of the same composition as in Examples 9 and 10 is prepared by use of a heated magnetic stirrer (Pyro-Magnestir, Lab-Line). Lecithin is dispersed in tripalmitate as described above. The heated aqueous phase is added to the melt. The dispersion is produced by stirring the mixture at a temperature of 80 ° C. for 30 minutes. The dispersion is allowed to cool at room temperature.
The average particle size after preparation (by volume) of the SLP dispersion measured by laser diffraction measurement (Malvern Master-sizer MS20) is 59.5 μm. The maximum particle size measurement is 250 μm. In contrast to high pressure homogenization and probe sonication, agitation produces relatively large particles in the micrometer size range.

マトリックス構成分の融点が高くなるにつれてSLPの平均粒度が大きくなる。 Example 12 : Effect of matrix constituents on average particle size of SLP
SLP comprising 10% matrix component, 1.2% soy lecithin (Lipoid S 100), 0.4% glycocholic acid, 2.25% glycerol and 0.01% thiomersal in 100% total distilled water A dispersion is prepared as described in Example 1. Five different matrix constituents are used: cetyl palmitate, which is a wax, and trilaurate, trimyristate and tripalmitate, which are white wax and triglyceride.
Table 2 shows the PCS average particle size and the melting point of the matrix constituents of various SLP dispersions.

The average particle size of SLP increases as the melting point of the matrix component increases.

Example 13 : Effect of Emulsifier Type and Amount on Average SLP Particle Size and Stability Tripalmitate SLP dispersions using various types and amounts of emulsifiers are prepared as described in Example 2. The composition of the various batches is shown in Table 3. All dispersions contain 2.25% glycerol and 0.01% thiomersal.
The average particle size of various batches of SLP is shown in FIG. The average particle size depends on the type and amount of emulsifier.

  The combination of phospholipids and bile salts is very efficient with respect to average particle size and stability. Systems stabilized only with phospholipids gel upon storage and form an ointment-like semi-solid gel. The system stabilized by Pluronic F68 shows a gelation tendency when shearing force is applied, that is, when the particles are forced to approach each other. Obviously, in this case, steric stabilization by the poloxamer is not sufficient. As a result, emulsifiers that are present on the lipid side and act from the lipid side (eg phospholipids) and emulsifiers in the dispersion medium that constitute highly mobile surfactant molecules (eg bile salts, tyloxapol and poloxamers) ) Surfactant combination is the optimum stabilization. Although repulsive force is an important factor for long-term stability, the basic mechanism of SLP stabilization is the high susceptibility of excess surfactant to immediately surface the surface to be regenerated during recrystallization of molten lipid. It is dynamic.

調製後にPCSにより測定された分散液の平均粒度を図13に示す。この実施例は、水性相中の共存乳化剤としての胆汁酸塩の燐脂質により安定化されたSLPに対する効果を実証するものである。胆汁酸塩を添加するとSLPの平均粒度を最高57％も下げることが明らかに示される。すなわち、胆汁酸塩を共存乳化剤として用いることにより、極めて微細な分散液を得ることができる。この胆汁酸塩の効果は、均質化過程で新たに生じる表面を界面活性剤分子が直ちに覆うことを可能にするミセル形成イオン性界面活性剤の高易動性に帰することができる。水性相中で液晶構造を形成する傾向のある燐脂質は新生分子を直ちに安定化される程易動性が十分でなく、そのため水性相中に高易動性の共存界面活性剤が存在しないと即座にコアレッセンスが起きてしまう。 Example 14 : Effect of bile salts as co-emulsifiers on SLP stabilized with phospholipids According to the method described in Example 1, bile salts (sodium glycocholate) were added or added to the aqueous phase Without preparation, SLP dispersions stabilized with phospholipids using various matrices (tripalmitate, hard fat) are prepared. All dispersions contain 2.25% glycerol and 0.01% thiomersal. The emulsification of the crude dispersion is performed by high pressure homogenization (APV Gaulin Micron Lab 40) under various homogenization conditions. The following dispersion was prepared.

The average particle size of the dispersion measured by PCS after preparation is shown in FIG. This example demonstrates the effect on BLP phospholipid stabilized SLP as co-emulsifier in aqueous phase. It is clearly shown that the addition of bile salts reduces the average particle size of SLP by as much as 57%. That is, a very fine dispersion can be obtained by using a bile salt as a co-emulsifier. This bile salt effect can be attributed to the high mobility of the micelle-forming ionic surfactant that allows the surfactant molecules to immediately cover the newly generated surface during the homogenization process. Phospholipids that tend to form a liquid crystal structure in the aqueous phase are not mobile enough to immediately stabilize the nascent molecule, so there must be a highly mobile coexisting surfactant in the aqueous phase. Coalescence occurs immediately.

Example 15 Preparation of Trimyristate SLP Stabilized by Lecithin / Bile Salt Blend 7.0 g of trimyristate (Dynasan 114, Huels AG) is melted at 70 ° C. in a thermostatic glass bottle. 0.96% phospholipid (Lipoid S 100) is dispersed in the melt by probe sonication. A solution of 280 mg sodium glycocholate, 1.6 g glycerol and 7 mg thiomersal in 61 ml of double distilled water is heated to 70 ° C. and added to the melt. A coarse dispersion is prepared by probe sonication for about 2 minutes. The crude dispersion is transferred to a high-pressure homogenizer (APV Gaulin Micron Lab 40) that has been incubated at about 90 ° C. and passed through the homogenizer five times at a pressure of 500 bar. The homogenized dispersion is allowed to cool at room temperature.
The average particle size after preparation measured by PCS is 155.7 nm. Laser diffraction measurement (Malvern Mastersizer MS20) cannot detect particles exceeding 1 μm. The particle size distribution obtained by laser diffraction measurement is shown in FIG. In this example, when a bile salt is used as a coexisting emulsifier of SLP stabilized by phospholipid, the formation of particles larger than 1 μm is effectively prevented by quickly covering the nascent surface during homogenization. This demonstrates that immediate coalescence is minimized.

Example 16 : Preparation of tripalmitate SLP without sonication (Method A)
Melt 4.0 g of tripalmitate (Dynasan 116, Huels AG) at 85 ° C. in a thermostatic glass bottle. 0.96 g of lecithin (Lipoid S 100) is dissolved in ethanol. The lecithin solution is added to the melt. Ethanol is evaporated at 85 ° C. 160 mg sodium glycocholate, 0.9 g glycerol and 4 mg thiomersal are dissolved in 35 ml double distilled water. The solution is heated to 85 ° C. and added to the melt. Prepare the crude dispersion by ultra-turrax vortexing for about 2 minutes. The crude dispersion is transferred to a high-pressure homogenizer (APV Gaulin Micron Lab 40) that has been incubated at about 90 ° C. and passed through the homogenizer 10 times at a pressure of 800 bar. The homogenized dispersion is allowed to cool at room temperature.

Example 17 : Preparation of tripalmitate SLP without sonication (Method B)
Melt 4.0 g of tripalmitate (Dynasan 116, Huels AG) at 85 ° C. in a thermostatic glass bottle. 0.96 g of lecithin (Lipoid S 100) is added to the melt. The mixture is shaken until the lecithin is completely dispersed in the melt and the dispersion is isotropic. 160 mg sodium glycocholate, 0.9 g glycerol and 4 mg thiomersal are dissolved in 35 ml double distilled water. The solution is heated to 85 ° C. and added to the melt. Prepare the crude dispersion by ultra-turrax vortexing for about 2 minutes. The crude dispersion is transferred to a high-pressure homogenizer (APV Gaulin MicronLab 40) that has been incubated at about 90 ° C. and passed through the homogenizer 10 times at a pressure of 800 bar. The homogenized dispersion is allowed to cool at room temperature.

Example 18 Preparation of Tripalmitate SLP by Dispersing Phospholipids in Aqueous Phase 4.0 g of tripalmitate (Dynasan 116) is melted at 80 ° C. in a thermostatic glass bottle. 0.96 g of phospholipid (Lipoid S 100) is dispersed in 35 ml of an aqueous solution of 160 mg sodium glycocholate, 0.9 g glycerol and 4 mg thiomersal by stirring for about 1 hour. The phospholipid dispersion is heated to 80 ° C. and added to the tripalmitate melt. A coarse dispersion is prepared by probe sonication for about 2 minutes. The crude dispersion is transferred to a high-pressure homogenizer (APV Gaulin Micron Lab 40) that has been incubated at about 90 ° C. and passed through the homogenizer 10 times at a pressure of 800 bar. The homogenized dispersion is allowed to cool at room temperature.

Example 19 Preparation of Tripalmitate SLP Stabilized with High Mobility Surfactant 5.0 g of tripalmitate is melted at 80 ° C. in a thermostatic glass bottle. 600 mg sodium glycocholate is dissolved in 44.4 g double-distilled water containing 1.13 g glycerol and 0.01% thiomersal. The aqueous solution is heated to 80 ° C. and added to the melt. A coarse dispersion is prepared by sonication for about 5 minutes. The crude dispersion is transferred to an isothermal high pressure homogenizer (APV Gaulin Micron Lab 40) and passed through the homogenizer 8 times at a pressure of 800 bar. The dispersion is allowed to cool at room temperature.
The average particle size (by number) of the prepared SLP dispersion measured by PCS is 96.8 nm. The particle size distribution is narrow and uniform.
This example is of one type, provided that the surfactant is highly mobile and constitutes an aqueous stabilizer reservoir to stabilize the nascent surface during recrystallization of the SLP matrix. It has been demonstrated that small, uniform particle size SLPs can be prepared by using only a surfactant such as sodium glycocholate, a bile salt.

これら分散液の平均粒度が18ケ月間の貯蔵の間、実際上不変のままであることがわかる。すなわち、これらの結果は、医薬不含のおよび医薬搭載SLP分散液が脂質乳濁液のそれに類似した長期安定性を示すことを実証している。 Example 20 : Long-term stability of various SLP dispersions Several different SLP dispersions are prepared according to the method described in Example 2. All dispersions contain 2.25% glycerol as an isotonic agent and 0.01% thiomersal as a preservative. The long-term stability of these dispersions is determined by repeated particle size measurements (by PCS) over a period of 18 months. These dispersions are stored at refrigeration temperatures. For comparison, a soybean oil emulsion system is included. The composition of these dispersions and their average particle size during storage are summarized in Table 4.

It can be seen that the average particle size of these dispersions remains practically unchanged during 18 months of storage. That is, these results demonstrate that drug-free and drug-loaded SLP dispersions exhibit long-term stability similar to that of lipid emulsions.

Example 21 : Sterile filtration of tripalmitate SLP 3% tripalmitate (Dynasan 116, Huels AG), 1.5% tyloxapol, 1% lecithin (Lipoid S 100), 2.25% glycerol and 0.01% thiomersal. 40 ml of a crude SLP dispersion comprising 100% total in double distilled water is prepared according to the method described in Example 6. The crude dispersion is passed 5 times through a constant temperature homogenizer (APV Micron Lab 40) at a pressure of 1200 bar. Half of the batch is allowed to cool at room temperature while the remainder is filtered through a sterile syringe filter (Nalgene SFCA, 0.2 μm pore size) before cooling to the recrystallization temperature of the molten lipid.
The particle size distribution of both samples is measured by PCS. The average particle size of the unfiltered sample is 56.7 nm and that of the sterile filtered SLP dispersion is 53.2 nm, i.e. both samples have practically the same particle size.

Example 22 : Sterilization of tripalmitate SLP by autoclaving 3% tripalmitate (Dynasan 116, Huels AG), 1.8% lecithin (Lipoid S 100), 0.6% sodium glycocholate, 2.25% glycerol and 40 ml of a crude SLP dispersion comprising 0.01% thiomersal in 100% total in double distilled water is prepared according to the method described in Example 2. The crude dispersion is passed 10 times through a constant temperature homogenizer (APV Micron Lab 40) at a pressure of 1200 bar.
The SLP dispersion is sterilized by filling into an injection glass bottle and autoclaving at 121 ° C / 2 atm for 45 minutes before cooling to the recrystallization temperature of the molten lipid. Allow the autoclaved dispersion to cool at room temperature. It shows no signs of aggregation or phase separation and has a PCS measured average particle size of 65.9 nm.

Example 23 : Lyophilization of SLP In a thermostatic glass bottle, 3.5 g of tripalmitate (Dynasan 116, Huels AG) is melted at 75 ° C. and 1.05 g of lecithin (Lipoid S 100, Lipoid KG) is melted. Disperse in the object by probe sonication. 1.58 g tyloxapol, 14 g sucrose and 7 mg thiomersal are dissolved in 50 ml double distilled water heated to 75 ° C. and the aqueous phase is added to the lipid melt. A crude dispersion is prepared by probe sonication and then passed 5 times through a constant temperature homogenizer (APV Gaulin Micron Lab 40) at a pressure of 900 bar. The homogenized dispersion is passed through a 0.2 μm sterilizing filter.
For lyophilization, the dispersion is filled into flat bottom glass bottles, the bottles are immersed in liquid nitrogen for 1 minute and transferred to a lyophilization chamber. Samples are lyophilized for 36 hours under vacuum at a recipient temperature of −40 ° C.
Freeze-drying yields a fine powder that can be easily redispersed. The particle size of the SLP dispersion is measured by PCS before lyophilization and after reconstitution of lyophilized powder with double distilled water. The average particle size before lyophilization is 79 nm and that of the reconstituted dispersion is 87 nm, ie there is virtually no change in the average particle size after lyophilization.

Example 24 : Surface modification by adsorption of polymer In a thermostatic glass bottle 4.0 g of tripalmitate (Dynasan 116, Huels AG) was melted at 80 ° C. and 1.6 g of soy lecithin (Lipoid S 100) in the melt. To be dispersed by probe sonication. 35.25 g of an aqueous solution of 0.01% thiomersal is heated to 80 ° C. and added to the melt. After producing a coarse dispersion by probe sonication, it is passed through a high-pressure homogenizer five times at a pressure of 1200 bar. The dispersion is filtered through a 0.2 μm syringe filter. Divide the batch into three equal parts. One part is diluted with the same amount of water and stored at 90 ° C. to prevent gelation of the phospholipid stabilized dispersion when cooled below the recrystallization temperature. The other two parts of the batch are each incubated overnight with an equal volume of 6% (w / w) poloxamer 407 (Pluronic F127, BASF) and 6% (w / w) poloxamine 908 (Tetronic 908, BASF) solution, In this case, the polymer solution is added to the SLP dispersion before being cooled to a temperature lower than the recrystallization temperature of SLP so that the polymer molecules can be used immediately as soon as a new surface is created for recrystallization. Both polymers are described in the literature as modifying the biodistribution of colloidal particles for intravenous administration.
The modification of the surface properties of tripalmitate SLP is demonstrated by the zeta potential difference. The zeta potential was measured by laser Doppler anemometry in a microelectrophoresis cell (Malvern Zetasizer 3). The results are summarized in Table 5.

SLPをポロキサマーおよびポロキサミン型のブロック共重合体とインキュベーションするとゼータ電位が減少する。ポリマーの吸着により表面はより疎水性となる。表面の疎水性は、RES(細網内皮系）活性およびコロイド粒子の生物内分布を支配する要因の一つであるとして記述されている。

Incubation of SLP with poloxamer and poloxamine type block copolymers decreases the zeta potential. The adsorption of the polymer makes the surface more hydrophobic. Surface hydrophobicity is described as one of the factors governing RES (reticuloendothelial system) activity and biodistribution of colloidal particles.

Example 25 : Preparation of SLPs loaded with the cardioprotectant covidadelenone Three different SLPs containing the cardioprotective drug coidecarenone are prepared. The composition of SLP is summarized in Table 6. All dispersions contain 2.25% glycerol and 0.01% thiomersal.
Batches 1 and 2 are prepared by dispersing lecithin in the molten matrix component as described above. Cobidecalenone is dissolved in this melt. After adding an aqueous phase containing sodium glycocholate, glycerol and thiomersal, a crude dispersion is prepared by probe sonication. It is transferred to a constant temperature homogenizer (APV Micron Lab 40) and passed 10 times through the homogenizer at a pressure of 800 bar. The dispersions are allowed to cool at room temperature.
Batch 3 is prepared by dispersing lecithin in the molten matrix component as described above. Ubidecarenone is dissolved in this melt. After adding an aqueous phase containing tyloxapol, glycerol and thiomersal, a crude dispersion is prepared by probe sonication. It is transferred to a constant temperature homogenizer (APV Micron Lab 40) and passed through the homogenizer 5 times at a pressure of 1200 bar. The dispersion is allowed to cool at room temperature.

Example 26 : Preparation of Oxazepam-loaded SLP Melt 7.0 g of tripalmitate at 80 ° C in a thermostatic glass bottle. 1.68 g lecithin and 140 mg oxazepam are dispersed in the melt by probe sonication. 60 ml of heated aqueous phase containing 280 mg sodium glycocholate, 1.58 g glycerol and 7 mg thiomersal is added to the melt and a crude dispersion is prepared by probe sonication. The crude dispersion is homogenized by passing 10 times through a constant temperature and high pressure homogenizer at a pressure of 800 bar. The dispersion is allowed to cool at room temperature. The dispersion of oxazepam-loaded SLP has an average particle size of 122.7 nm after preparation.

Example 27 : Preparation and long-term stability of diazepam-loaded SLP 4.0g of tripalmitate is melted at 80 ° C in a thermostatic glass bottle. 0.96 g lecithin and 120 mg diazepam are dispersed in the melt with the probe sonication. 35 ml of heated aqueous phase containing 160 mg sodium glycocholate, 0.9 g glycerol and 4 mg thiomersal is added to the melt and a crude dispersion is prepared by probe sonication. The crude dispersion is homogenized by passing 10 times through a constant temperature and high pressure homogenizer at a pressure of 800 bar. The dispersion is allowed to cool at room temperature.
This diazepam-loaded SLP dispersion has an average particle size after preparation of 104.6 nm. After 12 months storage, the average particle size measured by PCS is 113.9 nm. During storage, no precipitation of medicinal substances is detected macroscopically. The presence of medicinal crystals is not observed when the dispersion is investigated by polarization microscopy over a 12 month monitoring period.

Example 28 : Preparation of lidocaine-loaded SLP 4.0g of tripalmitate is melted at 80 ° C in a thermostatic glass bottle. 0.96 g lecithin and 60 mg lidocaine are dispersed in the melt by probe sonication. 35 ml of heated aqueous phase containing 320 mg sodium glycocholate, 0.9 g glycerol and 4 mg thiomersal is added to the melt and a crude dispersion is prepared by probe sonication. The crude dispersion is homogenized by passing 10 times through a constant temperature and high pressure homogenizer at a pressure of 800 bar. The dispersion is allowed to cool at room temperature.
This lidocaine loaded SLP dispersion has an average particle size after preparation of 90.4 nm.

Example 29 : Preparation and long-term stability of SLP loaded with prednisolone 4.0 g of tripalmitate is melted at 80 ° C in a thermostatic glass bottle. 0.48 g lecithin and 80 mg prednisolone are dispersed in the melt by probe sonication. 36 ml of heated aqueous phase containing 160 mg sodium glycocholate, 0.9 g glycerol and 4 mg thiomersal is added to the melt and a crude dispersion is prepared by probe sonication. The crude dispersion is homogenized by passing 10 times through a constant temperature and high pressure homogenizer at a pressure of 800 bar. The dispersion is allowed to cool at room temperature.
The dispersion of SLP with prednisolone has an average particle size after preparation of 118.3 nm. The average particle size measured by PCS after storage for 12 months is 124.2 nm. Material precipitation during storage is not detected macroscopically. Examination of the dispersion by polarized light microscopy over a 12 month monitoring period shows no presence of medicinal crystals.

Example 30 : Preparation of cortisone-loaded SLP Four different SLPs containing cortisone are prepared. The composition of these SLPs is summarized in Table 7. All dispersions contain 2.25% glycerol and 0.01% thiomersal.
Batches 1 and 2 are prepared by dispersing lecithin in the molten matrix component as described above. Cortisone is dissolved in this melt. After the addition of the aqueous phase containing sodium glycocholate, chrysolol and thiomersal, a crude dispersion is prepared by probe sonication. It is transferred to a constant temperature homogenizer (APV Micron Lab 40) and passed through the homogenizer 10 times. The dispersions are allowed to cool at room temperature.
Batch 3 is prepared by dispersing lecithin in the molten matrix component as described above. Cortisone is dissolved in this melt. After the addition of an aqueous phase containing poloxamer (Pluronic F 68), glycerol and thiomersal, a crude dispersion is prepared by probe sonication. It is transferred to a constant temperature homogenizer (APV Micron Lab 40) and passed through the homogenizer five times at a pressure of 1200 bar. The dispersion is allowed to cool at room temperature.
Batch 4 is prepared by dispersing lecithin in the molten matrix component as described above. Cortisone is dissolved in this melt. After the addition of the aqueous phase containing tyloxapol, glycerol and thiomersal, a crude dispersion is prepared by probe sonication. It is transferred to a constant temperature homogenizer (APV Micron Lab 40) and passed through the homogenizer 5 times at a pressure of 1200 bar. The dispersion is allowed to cool at room temperature.

Example 31 : Tripalmitate SLP with Retinol (Vitamin A)
Melt 1.0 g of tripalmitate (Dynasan 116, Huels AG) at 80 ° C. in a thermostatic glass bottle. 60 mg of retinol (vitamin A-alcohol> 99%, Fluka) is dissolved in the melt. 300 mg soy lecithin (Lipoid S 100) is dispersed in the melt by probe sonication until the dispersion is optically clear. 450 mg of poloxamer 407 (Pluronic ^™ F127, BASF) is dissolved in 29.0 g of double distilled water. The aqueous phase is heated to 80 ° C. and added to the melt. A fine dispersion is prepared by probe sonication for 20 minutes. The dispersion is filtered through a 0.2 μm syringe filter to remove spilled metal from the ultrasonic probe. The dispersion is allowed to cool at room temperature.
The average particle size by number after preparation of vitamin A-loaded tripalmitate SLP as measured by PCS is 98.5 nm.

Example 32 : Tripalmitate SLP with phytylmenadione (vitamin K ₃ )
Melt 1.0 g of tripalmitate (Dynasan 116, Huels AG) at 80 ° C. in a thermostatic glass bottle. 60 mg phytylmenadione (vitamin K ₃ , Sigma) and 300 mg soy lecithin (Lipoid S 100) are dispersed in the melt by probe sonication until the dispersion is optically clear. 450 mg of poloxamer 407 (Pluronic ^™ F127, BASF) is dissolved in 28.7 g of double distilled water. The aqueous phase is heated to 80 ° C. and added to the melt. A fine dispersion is prepared by probe sonication for 20 minutes. The dispersion is filtered through a 0.2 μm syringe filter to remove metal spilled from the ultrasonic probe. The dispersion is allowed to cool at room temperature. The average particle size by number after preparation of vitamin K ₃ -loaded tripalmitate SLP as measured by PCS is 86.8 nm.

Example 33 Preparation of Estramustine-Loaded Tripalmitate SLP 7.0 g of tripalmitate (Dynasan 116, Huels AG) is melted at 80 ° C. in a thermostatic glass bottle. 1.68 g of soy lecithin (Lipoid S100) is dispersed in the melt by probe sonication until the dispersion is optically clear. 40 mg estramustine is dissolved in the tripalmitate / lecithin dispersion. 0.42 g sodium glycocholate and 1.58 g glycerol are dissolved in 60 g double-distilled water. The aqueous phase is heated to 80 ° C. and added to the melt. A crude emulsion is prepared by probe sonication for about 2 minutes. The crude emulsion is transferred to a constant temperature high pressure homogenizer (APV Gaulin Micron Lab 40) and passed through the homogenizer 10 times at a pressure of 800 bar. The dispersion is allowed to cool at room temperature.

Example 34 : Physical state of various SLPs at body temperature Two batches of SLPs from different matrix components are prepared according to the method described in Example 2. Batch 1 consists of 10% tripalmitate, 0.5% ubidecarenone, 1.2% soy lecithin (Lipoid S 100, Lipoid KG), 0.4% sodium glycocholate, 2.25% glycerol, 0.01% thiomersal and 100% total % Is composed of double distilled water (% is weight). Batch 2 consists of 10% hard fat (Witepsol ^™ W35, Huels AG), 0.5% ubidecarenone, 1.2% soy lecithin (Lipoid S 100, Lipoid KG), 0.4% sodium glycocholate, 2.25% glycerol, 0.01 Consists of% thiomersal and double distilled water to 100% total (% is weight).
The physical state of these matrix components is measured at 20 ° C. and 38 ° C. by synchrotron radiation X-ray diffraction. Place the sample in a constant temperature sample holder. Each diffraction pattern is recorded for 180 seconds. FIG. 15a demonstrates that both batches of SLP are crystalline at room temperature (20 ° C.). Spacing corresponds to the β-crystal polymorph. At body temperature (38 ° C.), tripalmitate SLPs are still crystalline, whereas no reflexes are detected for hard fat SLPs, ie they are melted amorphous (FIG. 15b). The different physical states of these SLPs at body temperature result in different biopharmaceutical behavior with respect to the release of incorporated drugs or bioactive agents. SLP melted at body temperature basically exhibits the release characteristics typical of conventional lipid emulsions. Because the drug molecules diffuse freely in the liquid lipid, the drug can be released from the vehicle at a relatively high rate. In contrast, SLP that is solid at body temperature results in sustained release of the incorporated drug. Since the drug molecule is immobilized in the solid matrix, drug release is not controlled by diffusion but is dependent on the degradation of the solid lipid matrix in the body and is therefore delayed.

Example 35 : Preparation of PBA from miconazole 0.4 g of miconazole is melted at 90 ° C in a thermostatic glass bottle. 0.24 g of lecithin (Lipoid S 100) is dispersed in the melt by probe sonication until the dispersion is optically clear. 0.9 g glycerol, 80 mg sodium glycocholate and 4 mg thiomersal are dissolved in 38.5 ml double distilled water and heated to 90 ° C. The aqueous phase is added to the miconazole / lecithin melt and a coarse dispersion is produced by probe sonication for 5 minutes. The crude dispersion is transferred to a constant temperature high pressure homogenizer (APVGaulin Micron Lab 40) and passed 10 times through the homogenizer at a pressure of 800 bar. The PBA dispersion is allowed to cool at room temperature.
Upon cooling, the molten miconazole recrystallizes and forms a suspension of miconazole microparticles. The average particle size (by volume) of miconazole PBA measured by laser diffraction measurement is 21.8 μm. The miconazole PBA precipitate can be easily redispersed with light agitation.

Example 36 : Preparation of PBA from ibuprofen 1.2 g of ibuprofen is melted at 85 ° C in a thermostatic glass bottle. 0.72 g of lecithin (Lipoid S 100) is dispersed in the melt by probe sonication until the dispersion is optically clear. 0.9 g glycerol, 240 mg sodium glycocholate and 4 mg thiomersal are dissolved in 37 ml double distilled water and heated to 85 ° C. The aqueous phase is added to the ibuprofen / lecithin melt and a crude dispersion is produced by probe sonication for 5 minutes. The crude dispersion is transferred to a constant temperature high pressure homogenizer (APV Gaulin Micron Lab 40) and passed through the homogenizer 6 times at a pressure of 800 bar. The PBA dispersion is allowed to cool at room temperature.
Upon cooling, the molten ibuprofen recrystallizes and forms a suspension of ibuprofen microparticles. The average particle size (by volume) of ibuprofen PBA measured by laser diffraction measurement is 61.4 μm. The ibuprofen PBA precipitate can be easily redispersed with light agitation.

Example 37 : Dissolution rate of ibuprofen PBA The dissolution rate of ibuprofen PBA of Example 36 is measured with a laser diffractometer (Malvern Mastersizer MS20) by monitoring the decay of the so-called obscuration over 10 minutes. The degree of blurring is a measure of the unscattered laser light intensity drop by the sample and is related to the particle concentration in the laser beam. The particle size can be measured in parallel. For measurement, the ibuprofen PBA sample is diluted with water and dispersed by magnetic stirring in a measurement cell placed in the laser beam line. FIG. 16 shows the blurring attenuation and particle size of the ibuprofen PBA sample. Within 10 minutes, the degree of smearing decayed to zero, indicating that there was no detectable amount of particles, suggesting that PBA was completely dissolved. The dissolution of the raw raw material ibuprofen cannot be measured by this method because the wettability of the material in water is low.

Example 38 : Preparation of PBA from lidocaine 1.2g lidocaine is melted at 80 ° C in a thermostatic glass bottle. 1.2 g of tyloxapol is dissolved in 37.6 ml of double distilled water and heated to 80 ° C. The aqueous phase is added to the lidocaine melt and a crude dispersion is produced by probe sonication for 2 minutes. The crude dispersion is transferred to a constant temperature and high pressure homogenizer (APV Gaulin Micron Lab 40) and passed through the homogenizer five times at a pressure of 1200 bar. The PBA dispersion is allowed to cool at room temperature.
Upon cooling, the molten lidocaine recrystallizes into fine needles and forms a suspension of lidocaine particulates. FIG. 17 shows a polarization micrograph of the suspended lidocaine needles. As demonstrated by the polarization micrograph in FIG. 18, the particle shape of the raw lidocaine-base (Synopharm) is different from that of lidocaine PBA.
The average particle size (by volume) of lidocaine PBA measured by laser diffraction measurement is 174.2 μm. The maximum detection particle size is 400 μm. Lidocaine PBA sediments can be easily redispersed with light agitation. When water is added to PBA, the particles dissolve quickly. In contrast, raw material lidocaine is poorly soluble in water and has a much slower dissolution rate. This high dissolution rate of lidocaine PBA is a result of the modified surface properties and the finely dispersed state of the particles. Due to the rapid dissolution, it is not possible to measure the dissolution rate by the method described in Example 37.

Example 39 Preparation of PBA from Cholecalciferol (Vitamin D ₃ ) 0.8 g of cholecalciferol is melted at 95 ° C. in a thermostatic glass bottle. 120 mg soy lecithin (Lipoid S 100) is dispersed in the melt by probe sonication until the dispersion is optically clear. 40 mg sodium glycocholate and 0.9 g glycerol are dissolved in 37.92 ml double distilled water and heated to 95 ° C. The aqueous phase is added to the cholecalciferol / lecithin dispersion and a 5 minute probe sonication produces a crude dispersion. The crude dispersion is transferred to a constant temperature and high pressure homogenizer (APV Gaulin Micron Lab 40) and passed through the homogenizer 8 times at a pressure of 1200 bar. The PBA dispersion is allowed to cool at room temperature.
The average particle size after preparation according to the number of cholecalciferol PBA measured by PCS is 325.1 nm.

Example 40 : Preparation of PBA from estramustine 2 g estramustine is melted at 105 ° C in a thermostatic glass bottle. 0.8 g of soy lecithin (Lipoid S 100) is dispersed in the melt by probe sonication until the dispersion is optically clear. Dissolve 0.2 g sodium glycocholate and 0.9 g glycerol in 36.1 g double distilled water. The aqueous phase is heated to 95 ° C. and added to the melt. A crude emulsion is prepared by probe sonication for about 5 minutes. The crude emulsion is transferred to a constant temperature high pressure homogenizer (APV Gaulin Micron Lab 40) and passed 5 times through the homogenizer at a pressure of 1200 bar. The dispersion is allowed to cool at room temperature.

Transmission electron micrograph of tripalmitate SLP of Example 1 after storage at room temperature for 5 months. The bar represents 400 nm. Differential scanning calorimetry (DSC) thermogram of a) pure tripalmitate and b) tripalmitate SLP of Example 1. The transition peak corresponds to the melting of the β-crystal polymorph. FIG. 3 is a synchrotron radiation wide-angle X-ray diffraction pattern of the tripalmitate SLP of Example 2. FIG. The reflection corresponds to the β-crystal polymorph. 6 is a transmission electron micrograph of the unstable hard fat SLP of Example 5. FIG. The SLP dispersion gelled by forming a three-dimensional network during storage. The bar corresponds to 1000 nm. Particle size distribution of tripalmitate SLP of Example 1 after 15 months storage. The graph represents the polygonal PCS measurement result. Comparison with that of 10% tripalmitate PCS particle size distribution of the SLP dispersions with commercially available lipids emulsions Intraliped ^(R) 10%. The effect of microfluidization time on the average particle size of the hard fat SLP of Example 3. Storage stability of the hard fat SLP of Example 3 indicated by development of average particle size over the course of storage time (monitoring period: 12 months). The effect of homogenization pressure on the average particle size of tripalmitate SLP. Effect of the number of passes on homogenization on the average particle size of tripalmitate SLP (0 times corresponds to a coarse dispersion prepared by sonication). Effect of sonication time on average particle size of tripalmitate SLP prepared by probe sonication. Effect of emulsifier type and amount on the average particle size of tripalmitate SLP prepared according to Example 13. Effect of bile salt as co-emulsifier on average particle size of various phospholipid stabilized SLP dispersions of Example 14. The particle size distribution of the trimyristate SLP of Example 15 measured by laser diffraction measurement. Physical state of various drug-loaded SLPs a) at 20 ° C. and b) 38 ° C. measured by synchrotron radiation wide-angle X-ray diffraction. Dissolution rate of ibuprofen PBA of Example 37. 4 is a polarizing micrograph of lidocaine PBA of Example 38 (magnification: 150 ×). Polarized micrograph of the lidocaine raw material used in the production of lidocaine PBA (magnification: 150x).

Claims (22)

A method of emulsifying an insoluble or poorly water-soluble agent that is solid at room temperature selected from bioactive agents or pharmaceuticals,
a. Melting the agent,
b. Heating the dispersion medium to the same temperature as the molten agent;
c. One or more water-soluble or dispersible stabilizers that do not form a phase separation in the dispersion medium, and the amount of the stabilizer is sufficient to prevent particle aggregation after emulsification Add,
d. A crude dispersion is produced by sonication, high-speed stirring and / or hand shaking of the melted agent and dispersion medium in a liquid phase, and the dispersion is homogenized by high-pressure homogenization;
e. Allowing the homogenized dispersion to cool until solid particles are formed by recrystallization of the melted agent;
here,
Said stabilizers are natural and synthetic phospholipids, their hydrogenated derivatives and mixtures thereof, sphingolipids and glycosphingolipids; physiological bile salts; saturated and unsaturated fatty acids or fatty alcohols; ethoxylated fatty acids or fatty alcohols; Alkylaryl-polyether alcohols; esters and ethers of sugars or sugar alcohols with fatty acids or fatty alcohols; acetylated or ethoxylated mono- and diglycerides; synthetic biodegradable polymers; ethoxylated sorbitan esters Or sorbitan ether; and an amphiphilic compound that is an ionic and non-ionic surfactant selected from the group consisting of: amino acids, polypeptides and proteins;
A method characterized by performing the steps a to e.   The method according to claim 1, wherein one or more stabilizers soluble or dispersible in lipids are additionally added to the melted agent in the step c.   Choose a filter pore size that is large enough not to retain solid particles, and pass the homogenized dispersion through the filter to remove particulate contaminants before being cooled below the recrystallization temperature. The method according to claim 1 or 2, wherein:   Bioactive agents that exhibit low bioavailability and / or poor absorption from the intestine, have a melting point below 100 ° C., or that can be lowered to temperatures below 100 ° C. by the addition of adjuvants Or a medicinal anesthetic and narcotic selected from the group consisting of butanilicaine, fomocaine, isobutamben, lidocaine, lysokine, prilocaine, pseudococaine, tetracaine, trimecaine, tropopacaine and etomidate; methixene and profenamine Anticholinergic drugs; group consisting of alimenazine, vinedaline, perazine, chlorpromazine, fenpentadiol, phenanisole, fluanisole, mebenazine, methylphenidate, thioridazine, troxaton and trimipramine Antidepressants, psychostimulants and neuroleptics selected; an antiepileptic drug selected from the group consisting of dimethadione and nisetamide; selected from the group consisting of butconazole, chlorphenesine, etisazol, exesalamide, pecilosin and miconazole An antifungal agent; an anti-inflammatory agent selected from the group consisting of butibufen and ibuprofen; a bronchodilator that is bamifilin; A cardiovascular drug selected from the group consisting of nitrite, sloctide, triprolol, xibendol and biquidil; asperlin, chlorambucil, chlornafadine, mitotene, estra Cytostatic drug selected from the group consisting of mustine, taxol, penchromedin and trophosphamide; hyperemic drug selected from the group consisting of capsaitin and methylnicotinate; nicoclonate, oxprenolol, pilifibrate, simfibrate And a lipid lowering drug selected from the group consisting of thiadenol; an antispasmodic drug selected from the group consisting of aminopromazine, caroneline, diphemerin, phencarbamide, tiropramide and moxaverine; testosterone enanthate and testosterone- (4-methylpentanoate) A testosterone derivative selected from the group consisting of: azaperone and bramate; a psychotropic drug selected from the group consisting of azaperone and bramate; an allyldon virus quiescent; retinol, retinol acetate and Vitamin A derivatives selected from the group consisting of nor palmitate; vitamin E derivatives selected from the group consisting of tocopherol acetate, tocopherol succinate and tocopherol nicotinate; menadione; cholecalciferol; and acephate, cyfluthrin, azine phosphomethyl A pesticide, a pesticide and a herbicide selected from the group consisting of: Cypermethrin, substituted phenylthiophosphate, fenclophos, permethrin, piperonal, tetramethrin and trifluralin. The method of any one of these.   2. The method of claim 1, wherein the physiological bile salt is selected from the group consisting of sodium cholate, sodium dehydrocholate, sodium deoxycholate, sodium glycocholate and sodium taurocholate.   The method of claim 1 wherein the alkylaryl-polyether alcohol is tyloxapol.    The method according to claim 1, wherein the synthetic biodegradable polymer is a block copolymer of polyoxyethylene and polyoxypropylene oxide.    The method according to claim 1, wherein the protein is gelatin or albumin.   6. The method according to any one of claims 1 to 5, wherein the stabilizer is a combination of phospholipids and physiological bile salts.   10. The method of claim 9, wherein the phospholipid: physiological bile salt molar ratio is 2: 1 or higher.   11. A method according to claim 9 or 10, wherein the stabilizer is a combination of a phospholipid and sodium glycocholate in a molar ratio of 2: 1 to 4: 1.   12. The surface properties of the particles are modified after homogenization by polymer adsorption or by dialysis of a water-soluble surfactant to control the biodistribution of the particles. The method according to claim 1.   The method according to claim 1, wherein the dispersion is stirred with a magnetic stirrer during cooling.   The dispersion medium is a pharmacologically acceptable liquid selected from water, ethanol, propylene glycol, dimethyl sulfoxide or methyl isobutyl ketone, or a mixture thereof, which does not dissolve the agent. The method of any one of these.   The method according to claim 1, wherein the dispersion medium contains an isotonic agent and / or a cryoprotectant.   Dispersion medium is water-soluble or dispersible stabilizer; isotonic glycerol or xylitol; cryoprotectant sucrose, glucose, maltose or trehalose; electrolyte; buffer; anti-aggregation agent sodium citrate, sodium pyrophosphate or 16. The method of any one of claims 1 to 15, comprising sodium dodecyl sulfate; and one or more of preservatives.   The method according to any one of claims 1 to 16, characterized in that the dispersion is sterilized by autoclaving or sterile filtration before cooling to a temperature below the recrystallization temperature of the molten lipid.   In a subsequent step, the amount of the dispersion medium is reduced by evaporation, or removed by filtration, ultrafiltration or lyophilization to form liquid-free particles that can be reconstituted before use. 18. The method according to any one of items 17.   20. A suspension of bioactive agent particles (PBA) produced by the method of any one of claims 1-19, wherein the PBA exhibits low bioavailability and / or from the intestine. A bioactive agent or medicament that is poorly absorbed, has a melting point lower than 100 ° C. or can be lowered to a temperature lower than 100 ° C. by the addition of an adjuvant, butanilicine, fomocaine, isobutamben, lidocaine, lysokine, prilocaine, Anesthetics and narcotics selected from the group consisting of pseudococaine, tetracaine, trimecamine, tropopacaine and etomidate; anticholinergics selected from the group consisting of methixene and prophenamine; alimenazine, vinedaline, perazine, chlorpromazine, fenpenta Diol, phenanisole, fu Antidepressants, psychostimulants and neuroleptics selected from the group consisting of anisole, mebenazine, methylphenidate, thioridazine, troxatone and trimipramine; antiepileptics selected from the group consisting of dimethadione and nisetamide; butconazole, chlorphene An antifungal agent selected from the group consisting of syn, etizazole, exesaramide, pecilosin and miconazole; an anti-inflammatory agent selected from the group consisting of butibufen and ibuprofen; a bronchodilator bamifilin; Nicofibrate, penbutolol, pyrmenol, prenylamine, procainamide, propatyl nitrate, suloticidyl, triprolol, xibendol and biki A cardiovascular drug selected from the group consisting of asperrin, chlorambucil, chlornaphhazine, mitotene, estramustine, taxol, penchromedin and trophosphamide; a cytostatic drug selected from capsaitin and methylnicotinate Hyperemic drug selected from group; Nicosponate, oxprenolol, pyrifibrate, simfibrate and thiadenol lipid lowering drug; antispasmodic selected from the group consisting of aminopromazine, caroneline, diphemerin, phencarbamide, tiropramide and moxaverine A testosterone derivative selected from the group consisting of testosterone enanthate and testosterone- (4-methylpentanoate); a spermatozoon selected from the group consisting of azaperone and bramate Alliedone, a virus quiescent drug; Vitamin A derivative selected from the group consisting of retinol, retinol acetate and retinol palmitate; Vitamin E derivative selected from the group consisting of tocopherol acetate, tocopherol succinate and tocopherol nicotinate An insecticide, pesticide selected from the group consisting of acephate, cyfluthrin, azine phosphomethyl, cypermethrin, substituted phenylthiophosphate, fenclophos, permethrin, piperonal, tetramethrin and trifluralin; Suspension characterized by being a herbicide.   PBA is water insoluble at the emulsion preparation temperature and is not solubilized by excess stabilizer or forms micelles on its own, even before the PBA particle size cools to room temperature 20. Suspension of bioactive agent particles (PBA) according to claim 19, characterized in that they remain unchanged after cooling.   PBA may be partially water soluble at emulsion preparation temperature and / or form mixed micelles with excess stabilizer and / or form micelles on their own, resulting in dissolved organisms 20. The bioactive agent according to claim 19, wherein the particle size increases after cooling to room temperature by crystal growth and / or precipitation of the active agent and / or by mass transfer from small to large particles. Suspension of particles (PBA).   A liquid-free product produced by removing a dispersion medium from a suspension of bioactive agent particles (PBA) according to any one of claims 19 to 21 by filtration, ultrafiltration or lyophilization. particle.

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